Monthly sulfadoxine/pyrimethamine-amodiaquine or dihydroartemisinin-piperaquine as malaria chemoprevention in young Kenyan children with sickle cell anemia: A randomized controlled trial

Steve M Taylor; Sarah Korwa; Angie Wu; Cynthia L Green; Betsy Freedman; Sheila Clapp; Joseph Kipkoech Kirui; Wendy P O’Meara; Festus M Njuguna

doi:10.1371/journal.pmed.1004104

. 2022 Oct 10;19(10):e1004104. doi: 10.1371/journal.pmed.1004104

Monthly sulfadoxine/pyrimethamine-amodiaquine or dihydroartemisinin-piperaquine as malaria chemoprevention in young Kenyan children with sickle cell anemia: A randomized controlled trial

Steve M Taylor ^1,^2,^3,^*, Sarah Korwa ⁴, Angie Wu ², Cynthia L Green ^2,⁵, Betsy Freedman ¹, Sheila Clapp ², Joseph Kipkoech Kirui ⁴, Wendy P O’Meara ^1,³, Festus M Njuguna ^4,⁶

Editor: Lorenz von Seidlein⁷

PMCID: PMC9591057 PMID: 36215323

Abstract

Background

Children with sickle cell anemia (SCA) in areas of Africa with endemic malaria transmission are commonly prescribed malaria chemoprevention. Chemoprevention regimens vary between countries, and the comparative efficacy of prevention regimens is largely unknown.

Methods and findings

We enrolled Kenyan children aged 1 to 10 years with homozygous hemoglobin S (HbSS) in a randomized, open-label trial conducted between January 23, 2018, and December 15, 2020, in Homa Bay, Kenya. Children were assigned 1:1:1 to daily Proguanil (the standard of care), monthly sulfadoxine/pyrimethamine-amodiaquine (SP-AQ), or monthly dihydroartemisinin-piperaquine (DP) and followed monthly for 12 months. The primary outcome was the cumulative incidence of clinical malaria at 12 months, and the main secondary outcome was the cumulative incidence of painful events by self-report. Secondary outcomes included other parasitologic, hematologic, and general events. Negative binomial models were used to estimate incidence rate ratios (IRRs) per patient-year (PPY) at risk relative to Proguanil. The primary analytic population was the As-Treated population. A total of 246 children were randomized to daily Proguanil (n = 81), monthly SP-AQ (n = 83), or monthly DP (n = 82). Overall, 53.3% (n = 131) were boys and the mean age was 4.6 ± 2.5 years. The clinical malaria incidence was 0.04 episodes/PPY; relative to the daily Proguanil group, incidence rates were not significantly different in the monthly SP-AQ (IRR: 3.05, 95% confidence interval [CI]: 0.36 to 26.14; p = 0.39) and DP (IRR: 1.36, 95% CI: 0.21 to 8.85; p = 0.90) groups. Among secondary outcomes, relative to the daily Proguanil group, the incidence of painful events was not significantly different in the monthly SP-AQ and DP groups, while monthly DP was associated with a reduced rate of dactylitis (IRR: 0.47; 95% CI: 0.23 to 0.96; p = 0.038). The incidence of Plasmodium falciparum infection relative to daily Proguanil was similar in the monthly SP-AQ group (IRR 0.46; 95% CI: 0.17 to 1.20; p = 0.13) but reduced with monthly DP (IRR 0.21; 95% CI: 0.08 to 0.56; p = 0.002). Serious adverse events were common and distributed between groups, although compared to daily Proguanil (n = 2), more children died receiving monthly SP-AQ (n = 7; hazard ratio [HR] 5.44; 95% CI: 0.92 to 32.11; p = 0.064) but not DP (n = 1; HR 0.61; 95% CI 0.04 to 9.22; p = 0.89), although differences did not reach statistical significance for either SP-AQ or DP. Study limitations include the unexpectedly limited transmission of P. falciparum in the study setting, the high use of hydroxyurea, and the enhanced supportive care for trial participants, which may limit generalizability to higher-transmission settings where routine sickle cell care is more limited.

Conclusions

In this study with limited malaria transmission, malaria chemoprevention in Kenyan children with SCA with monthly SP-AQ or DP did not reduce clinical malaria, but DP was associated with reduced dactylitis and P. falciparum parasitization. Pragmatic studies of chemoprevention in higher malaria transmission settings are warranted.

Trial registration

clinicaltrials.gov (NCT03178643).

Pan-African Clinical Trials Registry: PACTR201707002371165.

In a randomized trial, Steve M Taylor and colleagues study the effects of 3 malaria chemoprevention regimens on clinical malaria incidence, P. falciparum infection, and dactylitis among children with sickle cell anemia in Kenya.

Author summary

Why was this study done?

Sickle cell anemia (SCA) is a very common condition among children born in malaria-endemic areas of sub-Saharan Africa, but their supportive care regimens are poorly tailored to African settings.
Among the complications that children with SCA suffer are more severe outcomes owing to malaria, and, therefore, many African countries recommend various malaria preventive regimens for children with SCA.
It is important to compare the efficacy and safety of these regimens in order to enhance the supportive care of African children with SCA.

What did the researchers do and find?

In this study, 3 malaria chemoprevention regimens were compared among children under 10 years old with SCA at a single site in Homa Bay, Kenya.
Children were randomly assigned to take daily Proguanil (which is the standard of care in Kenya), a monthly combination of sulfadoxine-pyrimethamine with amodiaquine (SP-AQ), or a monthly combination of dihydroartemisinin-piperaquine (DP), and then followed monthly for 12 months.
Cases of malaria were very low among all 3 groups, but the combination of DP reduced the risk of being infected by Plasmodium falciparum parasites and of dactylitis, which is a common complication of SCA.
DP was not associated with a higher rate of serious adverse events, but SP-AQ was associated with an unexpectedly higher rate of deaths that did not achieve statistical significance.

What do these findings mean?

Monthly DP may be an alternative to existing chemoprevention regimens for children with SCA owing to its safety, acceptability, and efficacy on hematologic events.
SP-AQ-associated mortality among children with SCA was unexpected and though it did not achieve statistical significance merits further evaluation.
Further studies are needed to compare chemoprevention regimens on parasitologic and hematologic outcomes in areas of high P. falciparum transmission and delivered through routine SCA providers.

Introduction

Sickle cell anemia (SCA) afflicts over 300,000 newborns annually [1], and most of these children will be born in malaria-endemic regions of sub-Saharan Africa. In African settings, the mortality rate of SCA children is 20 times that of non-SCA children [2], and up to 90% of newborns with SCA die before the age of 5 years [3]. Broadly, this severity is the result of both hematologic and infectious complications, including severe anemia, vaso-occlusive (painful) crises, bacteremia and bacterial pneumonias, and malaria. Additionally, many children lack access to routine supportive care for SCA, including early diagnosis, specialized follow-up, and provision of hydroxyurea and other prophylactic medications. A pressing need exists to design and implement comprehensive SCA care that is tailored to African settings.

Malaria is more severe and deadly in children with SCA [4,5] and can precipitate vaso-occlusive complications [6,7]. Therefore, many country-level guidelines recommend routine malaria chemoprevention, including daily Proguanil in Kenya [8] and Nigeria [9] and monthly sulfadoxine-pyrimethamine (SP) in Uganda [10]. These policies are supported by meta-analyses that suggest benefit [11,12], although there is a paucity of high-quality, comparative effectiveness studies of malaria chemoprevention regimens in African children with SCA.

We tested the efficacy of 3 chemoprevention regimens in Kenyan children with SCA: daily Proguanil, monthly sulfadoxine/pyrimethamine plus amodiaquine (SP-AQ), and monthly dihydroartemisinin-piperaquine (DP). Daily Proguanil is the standard of care in Kenya; monthly SP-AQ is efficacious in the African Sahel as Seasonal Malaria Chemoprevention [13]; and DP is highly effective as therapy and, in research studies, as monthly prevention in children [14] and pregnant women [15]. We compared these 2 alternate regimens to the standard of care on the efficacy to prevent clinical malaria and adverse hematologic outcomes among children aged 1 to 10 years with laboratory-confirmed SCA in Western Kenya.

Methods

Trial design and oversight

The Enhancing Preventive Therapy of Malaria in children with Sickle cell anemia in East Africa (EPiTOMISE) trial was an open-label, parallel assignment randomized trial. The trial protocol was approved by ethics committees of Moi University (0001907) and Duke University (Pro00077428), registered with clinicaltrials.gov (NCT03178643) and the Pan-African Clinical Trials Registry (PACTR201707002371165), and approved by the Kenyan Pharmacy and Poisons Board (ECCT/17/08/06). The Duke Clinical Research Institute oversaw the trial, with technical and infrastructural support from the Duke Global Health Institute and Moi University School of Medicine. Written informed consent in English, Kiswahili, or Dholuo was obtained from the parents or guardians of all children. Independent study monitoring was performed biannually; a medical safety monitor reviewed all severe adverse events (SAEs); and a Data Safety and Monitoring Board (DSMB) convened by the National Heart Lung and Blood Institute (NHLBI) provided biannual review of progress and safety. SP-AQ was supplied free of charge to the trial by Guilin Pharmaceuticals, which had no role in the design, conduct, analysis, reporting, or decision to report the results. This study is reported as per the Consolidated Standards of Reporting Trials (CONSORT) guideline (S1 Checklist).

Trial setting and population

EPiTOMISE was conducted in Homa Bay, Kenya, a town of historically high malaria transmission; in 2012 to 2013, 46% of children with fever presenting to Homa Bay County Hospital had malaria [16]. Children between 12 months and 10 years of age with homozygous hemoglobin S (HbSS) disease confirmed by hemoglobin electrophoresis and who met additional eligibility criteria were enrolled from a routine SCA clinic at Homa Bay County Hospital and randomized in 1:1:1 ratio to receive either daily Proguanil, monthly SP-AQ, or monthly DP in an open-label fashion. Randomization was performed through the electronic data capture program using a block allocation with a block size of 9. No stratification criteria were used.

The inclusion criteria for enrollment were as follows: age greater than 12 months and less than 10 years at enrollment; current attendance at or willingness to attend the study clinic; residence in either Homa Bay County or the Rongo or Awendo subcounties of Migori County; confirmed hemoglobin genotype of HbSS by electrophoresis or high-performance liquid chromatography (HPLC); no immediate, apparent, or reported plans to relocate residence in the next 2 years; ability to take oral medication and be willing to adhere to the medication regimen or caregiver willingness to give the medical regimen as prescribed; and ability and willingness of parent or legally authorized representative (LAR) to give informed consent. From May 2019 were added the criteria at screening of hemoglobin concentration ≥6.5 g/dL and alanine aminotransferase (ALT) ≤50 U/L. The exclusion criteria were as follows: taking routine antimalarial prophylaxis for another indication (including co-trimoxazole for HIV infection); known allergy or sensitivity to sulfadoxine, pyrimethamine, amodiaquine, proguanil, dihydroartemisinin, piperaquine, artemether, lumefantrine, penicillin (if under 5 years old), or derivatives of these compounds; known chronic medical condition other than SCA (i.e., malignancy, HIV) requiring frequent medical attention; currently participating in another clinical research study, or having participated in one in the prior 30 days; living in the same household as a previously enrolled study participant; chronic use of medications known to prolong the QT interval in children; Fridericia’s corrected QT interval (QTcF) >450 milliseconds (ms); or receipt of a transfusion of red blood cells in the 120 days prior to screening.

Trial interventions

Following randomization to chemoprevention regimen, children were dispensed standard doses of each study drug at monthly follow-up visits, which were self-administered by caregivers; dosing (see S1 Text) was weight-based for Proguanil and DP and age-based for SP-AQ. Both SP-AQ (SPAQ-CO) and DP (D-Artepp) were acquired from a manufacturer with WHO prequalification (Guilin), the former supplied without charge by the manufacturer; Proguanil was manufactured by Cosmos Ltd (Nairobi, Kenya). Per local guidelines, children under 5 years also received daily penicillin at standard age-based doses, and all children were enrolled in the Kenya National Hospital Insurance Fund. Adherence to administration of prevention regimen was assessed at each monthly follow-up visit by trial staff via structured queries.

Trial assessments

Caregivers were encouraged to call trial staff and report for an acute visit if the child was unwell. At these and at routine monthly visits, children with fever >38°C or a history of subjective or objective fever in the prior 24 hours were tested for malaria with a rapid diagnostic test (RDT) detecting the parasite antigens HRP2 and pLDH (SD Bioline Malaria Ag P.f/Pan COMBO, Alere). Children with positive RDT results were treated with a standard weight-based course of artemether-lumefantrine.

At each monthly visit, we performed a complete review of systems and physical exam, assessed concomitant medications, and collected an interval history, including interval tests or treatments for malaria with verification when possible by review of outside records, self-reported painful crises and dactylitis, and SAEs. Venous blood was collected every 3 months for complete blood count, serum creatinine (sCr), and ALT and every 6 months for hemoglobin electrophoresis; clinical laboratories met ISO 15189 requirements. From May 2019, hemoglobin concentration was also measured at each visit using a point-of-care test (HemoCue Hb 301). At each acute care and routine visit, capillary blood was collected as a dried blood spot (DBS) for post hoc molecular detection of Plasmodium falciparum. In children allocated to DP residing in Homa Bay town, we repeated an electrocardiogram (ECG) 4 to 6 hours following the third dose of each monthly DP course.

Outcome measures

All children were followed for 12 months after randomization. The primary outcome was clinical malaria incidence, defined as subjective or objective fever with the presence of P. falciparum parasites measured by a malaria RDT or by severe malaria using a standard definition [17]. Secondary parasitologic outcomes were hospitalization for malaria, light microscopy–positive malaria, unconfirmed malaria, and asymptomatic parasitization. The main hematologic outcome was the incidence of painful events, defined as an episode of pain lasting 2 hours or more without an obvious cause, and additional secondary hematologic outcomes were dactylitis, defined as pain or tenderness with or without swelling of the hands or feet, transfusion of blood products, severe anemia (defined as hemoglobin concentration <5.5 g/dL), and acute chest syndrome. See S1 Text for full outcome definitions. Safety outcomes included SAEs, ALT elevation, anemia, and, in DP recipients, prolongation of the QT interval corrected for heart rate by Fridericia’s method (QTcF). P. falciparum was detected in all DBS using a real-time PCR assay [18]. In selected participants, glucose-6-phosphate dehydrogenase (G6PD) was genotyped by PCR amplification and bidirectional Sanger sequencing across nucleotides 202 and 376 that encode the most common A- deficiency variant in East Africa [19,20], and polymorphisms in CYP2C8*2 were similarly genotyped (see S1 Text for molecular methods) [21,22].

Statistical analysis

To estimate the sample size, we assumed 3.7 episodes per patient-year (PPY) using the malaria rate in 2011 by children enrolled in the RTS, S/AS01 vaccine trial in Siaya, Kenya [23], which historically has had similar transmission to Homa Bay [24], and a similar expected distribution of episodes shifted by treatment effect factors. Using simulations, we found enrolling 65 patients per treatment arm would provide >90% power to detect a reduction of 40% in the DP arm, approximately 40% power to detect a reduction of 20% in the Proguanil arm, and >90% power to detect a reduction of 40% in either arm. Allowing for 20% loss to follow-up, a sample size of 246 (82 per group) provided at least 90% power, assuming that each comparison between experimental arm and control would be tested at the 0.0269 level (Dunnett’s correction) in order to preserve a nominal alpha level of 0.05.

The analysis was initially planned using the Intention-To-Treat (ITT) population. This was changed to the As-Treated (AT) population during the finalization of the statistical analysis plan owing to the unanticipated crossover from October 2018 to May 2019 from the SP-AQ group to Proguanil; this crossover resulted from the temporary suspension of SP-AQ administration at the request of the DSMB to allow assessment of 4 deaths among SP-AQ recipients. Participants who were still in active follow-up assigned to the SP-AQ arm in May 2019 were crossed back over to SP-AQ at that time. Analyses using the According-to-Protocol (ATP) were also evaluated to compare patients treated as randomized (e.g., did not crossover).

Results are presented by treatment arm using the mean, standard deviation (SD), median, 25th and 75th percentiles (Q1, Q3), and range as appropriate for continuous variables, as well as the count and percentage for non-missing data for categorical variables.

Primary and secondary incidence rates (IRs), or the number of episodes PPY at risk, for each experimental arm were compared to the control using a generalized regression model with a negative binomial (NB) distribution to allow for interdependence between multiple episodes. This model structure allowed the use of the actual follow-up time as the offset variable and a robust sandwich variance estimate. Model distributions considered included the NB, Poisson and zero-inflated Poisson, and NB with the distribution chosen based on the residual plots. Model results are presented using the incidence rate ratio (IRR) with 95% confidence interval (CI). Prespecified subgroup analyses explored the impact of changes to protocol and access to care prompted by the COVID-19 pandemic and other covariates on clinical malaria and painful events by including an interaction term in the model; the main pandemic-related protocol change was that monthly follow-up visits alternated between in-person and telephone visits. Additional covariates at enrollment included age, hydroxyurea use, sex, hospitalization in prior 12 months, and hemoglobin concentration. Secondary dichotomous outcomes are presented as cumulative incidence (CI) rates per treatment arm and compared to the control using Fine and Gray’s method to allow death to be a competing risk and censoring of patients lost to follow-up within the ITT and ATP populations. Comparisons are presented using hazard ratio (HR) with 95% CI. The AT population was not evaluated for comparing cumulative IRs due to multiple crossing of treatment arms with events possibly occurring in each crossed arm. S1 Analysis describes all comparisons of the monthly groups to the daily Proguanil group; analyses comparing monthly DP to monthly SP-AQ were added later.

Each primary and secondary outcome comparison used an α = 0.0269 per Dunnett’s test for multiple comparisons to the same control group. Tables were generated from complete case data only. No imputation of baseline or endpoint data was done. All analyses were conducted using SAS 9.4 (SAS Institute, Cary, NC, USA).

Results

Participants and follow-up

The trial was conducted between January 23, 2018, and December 15, 2020 (Fig 1A). A total of 315 children were screened and consented, and 246 were enrolled and randomized. Overall, 53.3% (n = 131) were boys; the mean age was 4.6 ± 2.5 years; 83.7% (n = 205) reported receiving routine sickle cell care; 69.0% (n = 169) reported being treated for malaria in the prior 12 months; and the mean hemoglobin concentration was 7.9 ± 1.2 g/dL. Baseline characteristics were similar across the 3 groups (Table 1). Between October 2018 and May 2019, the DSMB temporarily suspended administration of SP-AQ owing to the observation that the first 4 participant deaths occurred among children receiving SP-AQ. Pending the implementation of enhanced safety monitoring, addition of eligibility criteria, and adoption of a stopping rule, the SP-AQ group temporarily crossed over to Proguanil; SP-AQ dispensing resumed in May 2019. Overall, of 83 children randomized to SP-AQ, 31 (37.3%) crossed to Proguanil, a mean of 155 ± 82 days after randomization, and 17 of these (54.8%) remained on study in May 2019 and subsequently restarted SP-AQ. As a result of this unexpected protocol change, primary efficacy analyses were performed on the AT population, defined as the population of all participants with treatment assignment reflecting the actual treatment received during the month under observation, and safety analyses performed on the ATP population. Overall 8.1% (n = 20) of children were withdrawn or lost to follow-up.

Table 1. Characteristics of the patients at randomization.

	All patients (N = 246)	Daily Proguanil (N = 81)	Monthly SP-AQ (N = 83)	Monthly DP (N = 82)
Demographics
Boys, % (n/N)	53.3 (131/246)	45.7 (37/81)	56.6 (47/83)	57.3 (47/82)
Luo, % (n/N)	99.2 (244/246)	98.8 (80/81)	98.8 (82/83)	100 (82/82)
Age, year, mean (SD)	4.6 (2.5)	4.8 (2.6)	4.3 (2.3)	4.7 (2.6)
Mid-upper arm circumference, cm, mean (SD)^*	15.1 (1.2)	15.2 (1.2)	15.0 (1.1)	15.1 (1.3)
Medical history
Receiving regular sickle cell care, % (n/N)	83.7 (205/245)	79.0 (64/81)	90.2 (74/82)	81.7 (67/82)
Routine medications, % (n/N)
Proguanil	93.1 (229/246)	90.1 (73/81)	95.2 (79/83)	93.9 (77/82)
Folic acid	93.9 (231/246)	90.1 (73/81)	96.4 (80/83)	95.1 (78/82)
Penicillin^*	81.6 (102/125)	79.5 (31/39)	81.4 (35/43)	83.7 (36/43)
Hydroxyurea	45.5 (112/246)	51.9 (42/81)	44.6 (37/83)	40.2 (33/82)
Nights slept under an ITN in prior week, mean (SD)	6.9 (0.9)	7 (0.3)	6.7 (1.2)	6.9 (0.8)
House treated with IRS in prior 6 months, % (n/N)	26.6 (65/244)	33.3 (27/81)	21.0 (17/81)	25.6 (21/82)
Treated for malaria in prior 12 months, % (n/N)	69.0 (169/245)	71.6 (58/81)	65.9 (54/82)	69.5 (57/82)
Hospitalizations in prior 12 months, % (n/N)
None	50.2 (123/245)	51.9 (42/81)	52.4 (43/82)	46.3 (38/82)
1–2	43.3 (106/245)	39.5 (32/81)	42.7 (35/82)	47.6 (39/82)
3–10	6.1 (15/245)	7.4 (6/81)	4.9 (4/82)	6.1 (5/82)
>10	0.4 (1/245)	1.2 (1/81)	0	0
Pain crises in prior 12 months, % (n/N)
None	15.1 (37/245)	16.0 (13/81)	15.9 (13/82)	13.4 (11/82)
1–2	30.2 (74/245)	27.2 (22/81)	34.1 (28/82)	29.3 (24/82)
3–10	42.9 (105/245)	44.4 (36/81)	41.5 (34/82)	42.7 (35/82)
>10	11.8 (29/245)	12.3 (10/81)	8.5 (7/82)	14.6 (12/82)
Dactylitis episodes in prior 12 months, % (n/N)
None	51.0 (125/245)	50.6 (41/81)	50.0 (41/82)	52.4 (43/82)
Once	11.4 (28/245)	16.0 (13/81)	7.3 (6/82)	11.0 (9/82)
Twice	11.0 (27/245)	11.1 (9/81)	12.2 (10/82)	9.8 (8/82)
3–5	14.3 (35/245)	16.0 (13/81)	17.1 (14/82)	9.8 (8/82)
6–10	5.7 (14/245)	1.2 (1/81)	7.3 (6/82)	8.5 (7/82)
>10	6.5 (16/245)	4.9 (4/81)	6.1 (5/82)	8.5 (7/82)
Laboratory measures
Hemoglobin, g/dL, mean (SD)^**	7.9 (1.2)	7.9 (1.1)	7.6 (1.1)	8.0 (1.2)
% Hemoglobin F, median (Q1, Q3)	9.9 (4.4, 17.7)	9.3 (5.1, 17.7)	9.7 (4.4, 16.7)	12.0 (4.4, 19.3)
ALT, U/L, mean (SD)^**	20.9 (8.7)	21.4 (9.7)	20.8 (8.3)	20.5 (8.3)
sCr, umol/L, mean (SD)^**	21.5 (6.7)	22.3 (6.6)	20.7 (6.4)	21.6 (7.2)
QTcF interval, mean (range)	427 (308, 450)	430 (386, 450)	424 (308, 448)	426 (369, 450)

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*Only in those <5 years.

**Measured either during screening or at enrollment visit.

ALT, alanine aminotransferase; DP, dihydroartemisinin-piperaquine; g/dL, grams per deciliter; IRS, indoor residual spraying; ITN, insecticide-treated bed net; QTcF, QT interval corrected for heart rate using Fridericia’s method; Q1, Q3, 25th and 75th percentiles; sCr, serum creatinine; SD, standard deviation; SP-AQ, sulfadoxine-pyrimethamine-amodiaquine; U/L, units per liter.

Owing to substantial local malaria transmission, county authorities began a campaign in February 2018 of indoor residual spraying with primiphos-methyl. Each yearly campaign lasted 6 weeks and was reinitiated in January 2019 and February 2020 (Fig 1B).

Adherence to prescribed chemoprevention was overall high (Table A in S1 Text), although more guardians reported difficulty getting children to take SP-AQ (27.5%) than Proguanil (3.7%) or DP (6.3%) and forgetting to administer Proguanil (16%) than SP-AQ (2.5%) or DP (0%).

Efficacy

We recorded 9 primary outcome malaria events overall, yielding a clinical malaria event IR of 0.04 events/PPY at risk (Table 2). In the AT population, the rates were not significantly different to the Proguanil group (0.03/PPY) in both the recipients of SP-AQ (0.09/PPY; IRR: 3.05; 95% CI: 0.36 to 26.14; p = 0.39) and DP (0.04/PPY; IRR: 1.36; 95% CI: 0.21 to 8.85; p = 0.90). Similar malaria rates were observed in the ITT population (Table 2), and when comparing DP to SP-AQ recipients (Table B in S1 Text).

Table 2. Main parasitologic and hematologic outcomes.

	Overall (n = 246)	Daily Proguanil (n = 81)	Monthly SP-AQ (n = 83)	Monthly DP (n = 82)
AT population
Patient-years follow-up	208.52	82.5	56.7	69.3
Clinical malaria
Number of events	9	3	3	3
Incidence rate per patient-year (95% CI)	0.04 (0.02–0.08)	0.03 (0.01–0.09)	0.09 (0.02–0.39)	0.04 (0.01–0.13)
Incidence rate ratio (95% CI)^*			3.05 (0.36–26.14)	1.36 (0.21–8.85)
p-value			0.39	0.90
Painful events
Number of events	897	373	238	286
Incidence rate per patient-year (95% CI)	4.30 (3.77–4.2)	4.42 (3.63–5.38)	4.28 (3.38–5.42)	4.18 (3.19–5.46)
Incidence rate ratio (95% CI)^*			0.97 (0.70–1.35)	0.95 (0.65–1.38)
p-value			0.96	0.92
ITT population
Patient-years follow-up	208.5	71.6	67.6	69.3
Clinical malaria
Number of events	9	3	3	3
Incidence rate per patient-year (95% CI)	0.04 (0.02–0.08)	0.04 (0.01–0.13)	0.04 (0.01–0.14)	0.04 (0.01–0.13)
Incidence rate ratio (95% CI)^*			1.06 (0.18–6.28)	1.03 (0.17–6.12)
p-value			>0.99	>0.99
Painful events
Number of events	897	339	272	286
Incidence rate per patient-year (95% CI)	4.36 (3.81–4.98)	4.77 (3.85–5.92)	4.11 (3.29–5.13)	4.17 (3.22–5.40)
Incidence rate ratio (95% CI)^*			0.86 (0.61–1.22)	0.87 (0.60–1.28)
p-value			0.54	0.65

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*Relative to daily Proguanil.

AT, As-Treated; CI, confidence interval; DP, dihydroartemisinin-piperaquine; ITT, Intention-To-Treat; SP-AQ, sulfadoxine/pyrimethamine-amodiaquine.

We recorded 209 self-reported painful events, yielding a rate of 4.30 events/PPY at risk (Table 2). In the AT population, compared to the rate of painful events in the Proguanil group (4.42/PPY), painful events were not significantly different in the recipients of SP-AQ (4.28/PPY; IRR: 0.97; 95% CI: 0.70 to 1.35; p = 0.96) and DP (4.18/PPY; IRR: 0.95; 95% CI: 0.65 to 1.38; p = 0.92). Similar rates and differences were observed in the ITT population, and when comparing DP to SP-AQ recipients.

Among secondary parasitologic outcomes, DP reduced asymptomatic parasitization by P. falciparum by 79% (IRR: 0.21; 95% CI: 0.08 to 0.56; p = 0.002). Most other secondary parasitologic outcomes were rare and did not vary significantly between chemoprevention groups (Table 3); results in the ITT population were similar (Table C in S1 Text). Among secondary hematologic outcomes, we recorded 81 episodes of dactylitis (0.84 episodes/PPY), and the incidence of dactylitis was reduced relative to that in Proguanil recipients (1.08 episodes/PPY) in the recipients of DP (0.51/PPY; IRR: 0.47; 95% CI: 0.23 to 0.96; p = 0.038). Compared to DP recipients, SP-AQ recipients had similar incidences of asymptomatic P. falciparum parasitization (IRR 2.23; 95% CI: 0.68 to 7.29; p = 0.23) and dactylitis (IRR 1.66; 95% CI: 0.77 to 3.56; p = 0.24) (Table B).

Table 3. Secondary outcomes in the AT population.

	Overall (n = 246)	Daily Proguanil (n = 81)	Monthly SP-AQ (n = 83)	Monthly DP (n = 82)
Patient-years follow-up	208.52	82.5	56.7	69.3
Parasitologic
Severe malaria
Incidence rate per patient-year (95% CI)	0	0	0	0
Hospitalized for malaria
Number of events	7	2	2	3
Incidence rate per patient-year (95% CI)	0.04 (0.02–0.09)	0.02 (0–0.08)	0.02 (0.01–0.11)	0.08 (0.02–0.32)
Incidence rate ratio (95% CI)^*			1.34 (0.13–13.70)	4.17 (0.42–41.34)
p-value			0.94	0.27
Light microscopy–positive malaria
Number of events	47	18	10	19
Incidence rate per patient-year (95% CI)	0.19 (0.14–0.26)	0.18 (0.10–0.32)	0.15 (0.07–0.31)	0.25 (0.15–0.42)
Incidence rate ratio (95% CI)^*			0.80 (0.28–2.32)	1.35 (0.56–3.27)
p-value			0.85	0.65
Unconfirmed malaria
Number of events	107	46	21	40
Incidence rate per patient-year (95% CI)	0.50 (0.39–0.65)	0.49 (0.33–0.71)	0.45 (0.26–0.81)	0.57 (0.40–0.82)
Incidence rate ratio (95% CI)^*			0.93 (0.45–1.96)	1.18 (0.65–2.13)
p-value			0.97	0.76
Fatal malaria
Cumulative incidence rate (95% CI)	0	0	0	0
*Asymptomatic P. falciparum* infection**
Number of events	74	50	15	9
Incidence rate per patient-year (95% CI)	0.35 (0.24–0.50)	0.58 (0.36–0.96)	0.27 (0.14–0.52)	0.12 (0.06–0.25)
Incidence rate ratio (95% CI)^*			0.46 (0.17–1.20)	0.21 (0.08–0.56)
p-value			0.13	0.002
Hematologic
Dactylitis
Number of events count	180	98	45	37
Incidence rate per patient-year (95% CI)	0.84 (0.62–1.13)	1.08 (0.68–1.71)	0.84 (0.53–1.35)	0.51 (0.33–0.78)
Incidence rate ratio (95% CI)^*			0.78 (0.41–1.50)	0.47 (0.23–0.96)
p-value			0.60	0.04
Severe anemia ^*
Number of events	21	9	8	4
Incidence rate per patient-year (95% CI)	0.10 (0.07–0.15)	0.11 (0.06–0.21)	0.14 (0.07–0.29)	0.06 (0.02–0.16)
Incidence rate ratio (95% CI)^*			1.30 (0.43–3.91)	0.53 (0.14–2.07)
p-value			0.81	0.47
Transfusion of blood products
Number of events	24	15	5	4
Incidence rate per patient-year (95% CI)	0.15 (0.08–0.31)	0.18 (0.07–0.44)	0.09 (0.03–0.30)	0.19 (0.04–0.99)
Incidence rate ratio (95% CI)^*			0.52 (0.10–2.79)	1.08 (0.13–9.19)
p-value			0.58	>0.99
Acute chest syndrome
Number of events	4	1	3	0
Incidence rate per patient-year (95% CI)	0.01 (0–0.05)	0.01 (0–0.09)	0.05 (0.01–0.20)	0
Incidence rate ratio (95% CI)^*			3.96 (0.27–57.42)
p-value			0.37
General
All-cause hospitalization
Number of events	139	56	43	40
Incidence rate per patient-year (95% CI)	0.67 (0.54–0.83)	0.70 (0.49–0.99)	1.28 (0.54–2.99)	0.78 (0.44–1.37)
Incidence rate ratio (95% CI)^*			1.83 (0.63–5.31)	1.11 (0.52–2.39)
p-value			0.35	0.93
Death
Cumulative incidence	10	2	7	1
Cumulative incidence rate (95% CI)	4.4% (2.4–8.1)	2.2% (0.5–9.3)	11.3% (6.1–20.7)	1.3% (0.2–9.1)
Hazard ratio (95% CI)^*			5.44 (0.92–32.11)	0.61 (0.04–9.22)
p-value			0.064	0.89

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*Defined as hemoglobin concentration <5.5 g/dL.

AT, As-Treated; CI, confidence interval; DP, dihydroartemisinin-piperaquine; g/dL, grams per deciliter; SP-AQ, sulfadoxine/pyrimethamine-amodiaquine.

In the AT population analyses of prespecified subgroups on main hematologic outcomes, rates of painful events did not vary between chemoprevention recipients within groups (Fig 2, Table D in S1 Text). These subgroups also did not modify the association of DP with reduced rates of dactylitis, which was observed irrespective of age, sex, baseline hemoglobin, or use of hydroxyurea.

Fig 2 — Points are estimates of the IRR relative to daily Proguanil. Bars are 95% CI. AT, As-Treated; CI, confidence interval; DP, dihydroartemisinin-piperaquine; g/dL, grams per deciliter; IRR, incidence rate ratio; SP-AQ, sulfadoxine/pyrimethamine-amodiaquine.

Safety

There were 102 SAEs overall in the AT population, in whom 41.9% (90/215) experienced at least 1 SAE, yielding an IR of 0.91 SAEs/PPY at risk. The incidence of SAEs was similar in recipients of Proguanil (0.90 events/PPY; 95% CI: 0.64 to 1.26), SP-AQ (1.10/PPY; 95% CI: 0.78 to 1.54), and DP (0.82/PPY; 95% CI: 0.54 to 1.24) (Table 4). The most commonly reported SAEs were hospitalizations resulting from a painful crisis (91 events), from anemia (33 events), and malaria or sepsis (12 events each) (Table E in S1 Text). One SAE was judged by the safety monitor to be related to chemoprevention regimen: an ALT elevation to 517 in a recipient of SP-AQ, which normalized following cessation of drug. We recorded 72 all-cause hospitalizations in the AT population, yielding an overall rate of hospitalization of 0.67/PPY at risk (Table 4); the prevalence and the rate of hospitalization was similar between groups. Ten children died: 2 receiving daily Proguanil, 7 receiving SP-AQ, and 1 receiving DP (Table 4); relative to Proguanil (cumulative IR [CIR] 2.2%), the risk of death was elevated in SP-AQ recipients (CIR 11.3%; HR 5.44; 95% CI 0.92 to 32.11; p = 0.064), although this did not reach statistical significance and was not elevated in DP recipients (CIR 1.3%; HR 0.61; 95% CI 0.04 to 9.22; p = 0.89). Similarly, relative to DP recipients, the risk of death was elevated in SP-AQ recipients (HR 8.86; 95% CI: 0.74 to 106.55; p = 0.10), although this did not achieve statistical significance (Table B in S1 Text).

Table 4. Safety outcomes and SAEs in ATP population.

	Overall (n = 215)	Daily Proguanil (n = 81)	Monthly SP-AQ (n = 52)	Monthly DP (n = 82)
General outcomes
All-cause hospitalizations, % (n)	33.5 (72)	34.6 (28)	36.5 (19)	30.5 (25)
Hospitalization incidence, events/PPY (95% CI)	0.68 (0.54–0.86)	0.66 (0.46–0.94)	0.85 (0.59–1.23)	0.60 (0.38–0.95)
Study drug withdrawn, % (n)	0.5 (1)	0	1.9 (1)	0
All-cause deaths, % (n)^**	4.2 (9)	2.5 (2)	11.5 (6)	1.2 (1)
Any SAE, % (n)	41.9 (90)	44.4 (36)	48.1 (25)	35.4 (29)
SAE incidence, events/PPY (95% CI)	0.91 (0.74–1.13)	0.90 (0.64–1.26)	1.10 (0.78–1.54)	0.82 (0.54–1.24)
Laboratory outcomes
Change in Hb from baseline to 12 months, mean (range), g/dL	0.2 (−2.5, 3.6)	0 (−2.5, 2.2)	0.4 (−2.1, 3.6)	0.3 (−1.8, 3.5)
Drop in Hb >2 g/dL from baseline, % (n)	6.1 (13)	3.7 (3)	3.9 (2)	9.8 (8)
Change in creatinine from baseline to 12 months, mean (range), umol/L	0.78 (−25.7, 19.4)	−0.31 (−25.7, 18.6)	0.50 (−13.6, 13.1)	2.05 (−18.8, 19.4)
sCr >62, % (n)	0.5 (1)	0	2.0 (1)	0
Change in ALT from baseline to 12 months, mean (range), U/L	5.39 (−66.2, 605.1)	8.64 (−66.2, 605.1)	4.38 (−23.4, 72.5)	2.55 (−37.5, 97.4)
Serum ALT >60, % (n)	7.5 (16)	8.6 (7)	5.9 (3)	7.3 (6)
ECG outcomes^*
Any QTcF >450 ms, % (n/N)	NA	NA	NA	50.0 (6/12)
Any QTcF change from baseline >50 ms, % (n/N)^*	NA	NA	NA	8.3 (1/12)

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*Among only monthly DP recipients enrolled in the monthly ECG monitoring substudy.

**One additional participant allocated to SP-AQ died but does not appear in the ATP population owing to their crossover to the Proguanil arm.

ALT, alanine aminotransferase; ATP, According-to-Protocol; CI, confidence interval; DP, dihydroartemisinin-piperaquine; ECG, electrocardiogram; Hb, hemoglobin; ms, millisecond; PPY, per patient-year; QTcF, QT interval corrected for heart rate using Fridericia’s method; SAE, serious adverse event; sCr, serum creatinine; SP-AQ, sulfadoxine/pyrimethamine-amodiaquine; U/L, units per liter.

Among laboratory events, neither the change in hemoglobin concentration from baseline nor the incidence of severe anemia, defined as a drop of more than 2 g/dL from baseline, differed between chemoprevention groups (Table 4). Similarly, there were no differences between groups in the change in baseline nor the incidence of elevated values of ALT or of sCr. Follow-up neutrophil and platelet counts in the ITT population are presented in Tables F and G in S1 Text. Among the 7 children who died while receiving SP-AQ, 2 were G6PD deficient (2 hemizygote boys), and 1 was heterozygote for a CYP2C8*2 allele associated with reduced metabolism of AQ (Table H in S1 Text).

Among DP recipients, we enrolled 12 children in a substudy of QTcF measurements following the third dose of their monthly courses of DP. Among 134 follow-up QTcF measurements in these children, we recorded 16 (11.9%) measurements of QTcF >450 ms, which occurred in 50% (6/12) of children, with a maximum QTcF of 479 ms (Figure A in S1 Text). There was 1 (0.7%) case of a prolongation of >50 ms QTcF compared to baseline to 52 ms (Table 4, Table I in S1 Text). Mean (SD) QTcF changes from baseline peaked at 17 (13) ms at 6 months and then declined to 8 (16) ms after 12 months of administration.

Discussion

In this randomized, open-label trial, we compared 2 monthly malaria chemoprevention regimens with the standard-of-care of daily Proguanil in Kenyan children with SCA in an historically malaria-holoendemic setting in Western Kenya. Consistent with enhanced community malaria control that sharply reduced transmission, we recorded very few episodes of malaria. Compared to Proguanil, neither monthly SP-AQ nor DP reduced the incidence of the primary outcome clinical malaria, although monthly DP did significantly reduce the incidence of asymptomatic parasitization. Hematologic outcomes were common, and we observed a significant reduction in dactylitis in the children receiving monthly DP. Given the observed safety of DP, the high adherence to it in our participants, and its known high efficacy as malaria prevention, monthly DP is a promising and feasible alternative as malaria chemoprevention in children with SCA to prevent both parasitologic and hematologic morbidity.

Monthly DP was associated with a reduced rate of episodes of dactylitis compared to daily Proguanil, both overall and in each examined subgroup. The prophylactic combination of penicillin and chloroquine reduced dactylitis in an early Ugandan trial [25], contributing to current recommendations for malaria chemoprophylaxis [11]. Monthly DP is highly effective as malaria prevention in children and in pregnant women owing to its immediate parasiticidal effect as well as the prolonged half-life of the piperaquine component, and, consistent with this, we observed a nearly 80% reduction in asymptomatic P. falciparum infections in DP recipients. Notably, this effect of DP on dactylitis was not observed on painful events, despite the more than 5-fold increase in the overall incidence of painful events and presumably similar pathophysiology. Other measured hematologic events were too rare to capture differences between groups, and we did not observe a difference in the change in hemoglobin concentration between groups. Whether a measurable effect of malaria chemoprophylaxis on common hematologic outcomes like painful crises can be captured in a higher-transmission setting remains an open question.

We observed for DP high adherence and acceptability by children (Table A in S1 Text). We recorded no drug-related SAEs for DP, and the scope and incidence of SAEs were similar to that for SP-AQ and Proguanil. As expected, we observed a prolongation of the QTcF interval following the third dose after repeated courses in a small subset of recipients. Although half of monitored children had at least 1 QTcF exceeding 450 ms, in only 1 recording was the QTcF prolonged beyond 50 ms from baseline, and no recipient required DP alteration. It is notable that 21 children failed screening owing to QTcF >450 ms, highlighting that baseline QT prolongation is common in these children. This may merit baseline QT measurement to limit DP-induced prolongation, although this risk may be mitigated by alternate DP dosing schemes or by, as we observed, the plateauing of QT intervals after 6 months of prophylaxis (Table I and Figure A in S1 Text). These clinical factors would need to be complemented before wider policy adoption by considerations of DP resistance, which was detected as declining clinical efficacy first in Southeast Asia [26] and more recently at several sites in Africa [27,28]. Although wider use of DP will necessarily increase pressure to develop resistance, this may be mitigated by a restriction of DP use to high-risk populations like pregnant women and children with SCA, and by the observation through multiple studies [29–32] that mass drug administration with DP across diverse populations has not reliably increased molecular markers of DP resistance.

On balance of efficacy and safety, monthly SP-AQ does not appear to have a role as chemoprevention in children with SCA. Reductions in asymptomatic P. falciparum infection and dactylitis did not reach statistical significance, and while these were undermined by low malaria transmission and likely by prevalent resistance to SP in Kenya [33], rates of general outcomes did not suggest benefit of the putative antibacterial activity of the sulfadoxine component. More notable is the severity of SAEs with SP-AQ. Seven children receiving SP-AQ died, and 1 additional child required cessation owing to transaminitis. Causes of death in children receiving SP-AQ were variable, including severe anemia, acute chest syndrome, and painful crisis, which were among the most common SAEs overall, and the durations of SP-AQ treatment prior to death varied from 81 to 294 days (Table H in S1 Text). We investigated as a contributing factor the possibility of G6PD-linked anemia owing to the known risk following SP exposure [34,35], but the A- allele of G6PD deficiency was present in only 2 of these 7 children. This high rate of SAEs was unexpected given the favorable safety profile of monthly SP-AQ in recipients as Seasonal Malaria Chemoprophylaxis [36], during which 3 to 4 monthly courses are administered to all children under 5 years of age. This difference could be the result of our use of SP-AQ in a chronically ill population or accumulated risk following repeated monthly dosing.

Our trial had several limitations. As noted above, the low incidence of P. falciparum infection prevents meaningful assessment of the primary outcome. Given the plausible link between infection and hematologic outcomes, this may have also limited our ability to fully assess other outcomes. The temporary cross-over of SP-AQ recipients to Proguanil necessitated analytic changes and limited approaches to counting outcomes, and this was mitigated by prioritizing the AT population. Finally, generalizability may be limited by the common baseline use of hydroxyurea and, as a result of trial participation, the enhanced supportive care of participants. Pragmatic studies of prophylaxis delivered through routine sickle cell care can measure effectiveness in diverse care and transmission settings.

In summary, among Kenyan children under 10 years with SCA, monthly DP was very acceptable and reduced the incidence of dactylitis and asymptomatic P. falciparum infection despite very limited malaria transmission or clinical malaria. Monthly SP-AQ had no measurable benefit compared to daily Proguanil, and we observed a greater number of deaths in this group, although mortality differences were not statistically significant. Our results provide a rationale to consider the wider use of DP with appropriate monitoring as a routine component of care for children with SCA in malaria-endemic settings.

Supporting information

S1 CONSORT Checklist. CONSORT checklist.

(DOC)

Click here for additional data file.^{(218KB, doc)}

S1 Protocol. Study protocol.

(DOCX)

Click here for additional data file.^{(711.2KB, docx)}

S1 Analysis. Statistical analysis plan.

(DOCX)

Click here for additional data file.^{(54.6KB, docx)}

S1 Text. Supporting information.

Supplemental methods, figure, and tables.

(DOCX)

Click here for additional data file.^{(98.4KB, docx)}

Acknowledgments

We thank the children and their parents for their participation; the site staff Ernest Ojwang, Seline Miruni, Faith Ogolla, and Erick Ayaye; the trial pharmacists Collins Saina and Linet Kugo; data managers Steven Karuru and Edna Sang; the members of the data and safety monitoring board (David Ayuku, Walter Dzik, Gregory Kato, Karen Kessler, Jennifer Knight-Madden, Irene Marete, Rebecca Pentz, Jennifer Rothman, and Liz Turner); the members of our Community Advisory Board; Stanley Odanga and Terry Odero for community engagement; Stacey Gondi for external trial monitoring; John Humphrey for independent safety monitoring; the CEOs of Homa Bay County Hospital Drs. Lillian Kochola and Meshack Liru for their accommodation of trial activities; Gayle Passmore, Minal Bhojai, Varsha Gajjar, Casey Silver, Lucy Abel, George Ambani, Tabitha Jepkurgat, Debbie Drosdick, and Assumpta Nantume for their operational assistance; Laura Edwards and Sharon Stroud for statistical programming support; the AMPATH Reference Laboratory for assay support; and Francis Kithuku, Dominique Cole, Robert Rono, and Benta Kamire for their administrative support.

Abbreviations

ALT: alanine aminotransferase
AT: As-Treated
ATP: According-to-Protocol
CI: confidence interval
CIR: cumulative IR
DBS: dried blood spot
DP: dihydroartemisinin-piperaquine
DSMB: Data Safety and Monitoring Board
ECG: electrocardiogram
EPiTOMISE: Enhancing Preventive Therapy of Malaria in children with Sickle cell anemia in East Africa
G6PD: glucose-6-phosphate dehydrogenase
HbSS: homozygous hemoglobin S
HPLC: high-performance liquid chromatography
HR: hazard ratio
IR: incidence rate
IRR: incidence rate ratio
ITT: Intention-To-Treat
LAR: legally authorized representative
ms: millisecond
NB: negative binomial
NHLBI: National Heart Lung and Blood Institute
PPY: per patient-year
QTcF: Fridericia’s corrected QT interval
RDT: rapid diagnostic test
SAE: severe adverse event
SCA: sickle cell anemia
sCr: serum creatinine
SD: standard deviation
SP-AQ: sulfadoxine/pyrimethamine-amodiaquine

Data Availability

Data cannot be shared publicly because the signed informed consent form placed restrictions on sharing broadly. For inquiries of data use that fall within the scope of the study and therefore the consent authorization, please contact the study team at epitomise-study@duke.edu.

Funding Statement

Supported by the NHLBI (www.nhlbi.nih.gov) of the NIH (R01 HL134211 to SMT). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. SP-AQ was supplied free of charge to the trial by Guilin Pharmaceuticals, which had no role in the design, conduct, analysis, reporting, or decision to report the results.

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PLoS Med. doi: 10.1371/journal.pmed.1004104.r001

Decision Letter 0

Richard Turner

16 Mar 2022

Dear Dr Taylor,

Thank you for submitting your manuscript entitled "Monthly sulfadoxine/pyrimethamine-amodiaquine or dihydroartemisinin-piperaquine as malaria chemoprevention in young children with sickle cell anemia: A randomized controlled trial" for consideration by PLOS Medicine.

Your manuscript has now been evaluated by the PLOS Medicine editorial staff and I am writing to let you know that we would like to send your submission out for external assessment.

However, we first need you to complete your submission by providing the metadata that is required for full assessment. To this end, please login to Editorial Manager where you will find the paper in the 'Submissions Needing Revisions' folder on your homepage. Please click 'Revise Submission' from the Action Links and complete all additional questions in the submission questionnaire.

Please re-submit your manuscript within two working days, i.e. by Mar 18 2022 11:59PM.

Once your full submission is complete, your paper will undergo a series of checks in preparation for external assessment.

Feel free to email us at plosmedicine@plos.org if you have any queries relating to your submission.

Kind regards,

Richard Turner, PhD

Senior Editor, PLOS Medicine

rturner@plos.org

PLoS Med. doi: 10.1371/journal.pmed.1004104.r002

Decision Letter 1

Richard Turner

12 Apr 2022

Dear Dr. Taylor,

Thank you very much for submitting your manuscript "Monthly sulfadoxine/pyrimethamine-amodiaquine or dihydroartemisinin-piperaquine as malaria chemoprevention in young children with sickle cell anemia: A randomized controlled trial" (PMEDICINE-D-22-00862R1) for consideration at PLOS Medicine.

Your paper was discussed with an academic editor with relevant expertise and sent to independent reviewers, including a statistical reviewer. The reviews are appended at the bottom of this email and any accompanying reviewer attachments can be seen via the link below:

[LINK]

In light of these reviews, we will not be able to accept the manuscript for publication in the journal in its current form, but we would like to invite you to submit a revised version that addresses the reviewers' and editors' comments fully. You will appreciate that we cannot make a decision about publication until we have seen the revised manuscript and your response, and we expect to seek re-review by one or more of the reviewers.

In revising the manuscript for further consideration, your revisions should address the specific points made by each reviewer and the editors. Please also check the guidelines for revised papers at http://journals.plos.org/plosmedicine/s/revising-your-manuscript for any that apply to your paper. In your rebuttal letter you should indicate your response to the reviewers' and editors' comments, the changes you have made in the manuscript, and include either an excerpt of the revised text or the location (eg: page and line number) where each change can be found. Please submit a clean version of the paper as the main article file; a version with changes marked should be uploaded as a marked up manuscript.

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Please let me know if you have any questions, and we look forward to receiving your revised manuscript.

Sincerely,

Richard Turner, PhD

Senior editor, PLOS Medicine

rturner@plos.org

-----------------------------------------------------------

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Comments from academic editor:

The problem of underpowered malaria trials in the presence of malaria elimination efforts is no longer unusual. The efficacy of antimalarial prophylaxis against asymptomatic infections becomes a useful surrogate marker in such circumstances. The impact of the 3 drug regimens on asymptomatic infections should be included in the abstract. The authors could also create an additional endpoint by combining asymptomatic and symptomatic Pf infections to increase the power. The incidence rate of asymptomatic infections in children receiving proguanil (0.58/year)is halved by prophylaxis with SP-AQ (0.27/year) and this rate is further halved by DP (0.12/year) – table 3. Similarly, dactylitis decrease from 1.08/year in patients treated with proguanil to 0.84/yea in patients treated with SP-AQ and 0.51/year in patients in DP. There is an intriguing trend suggesting that SP-AQ is more beneficial than proguanil but less beneficial than DP. The authors conclude that there is no role for SP-AQ and DP is the solution. This may be jumping to conclusions as Reviewer 2 points out. SP-AQ has by now been used in 10s of millions of children on seasonal chemoprophylaxis we know it is safe, available and accepted. DP may be ideal, but policymakers may be reluctant to roll out DP for prophylaxis. The experience in Cambodia shows that DP -resistance is a real danger. If DP no longer works, there are few options left. This threat of emerging and spreading resistance should be discussed by authors.

The authors describe and discuss safety issues extensively but not in a helpful fashion. The excess mortality in the SP-AQ arm is not statistically significant this is most probably due to chance. The authors have to be careful not to mislead the reader. Similarly, the observed variability in QTc intervals sounds worrying the way it is described in the current text and was immediately picked up by reviewer 4. It is most unfortunate that the investigators only measured the QTc in the patients receiving DP. Without a control it is not possible to infer any causality. The variability in QTc has to be analysed much more carefully. At a minimum the authors have to show the QTc variability over the follow-up period for the 12 trial participants on whom they have serial QTcs. Leaving the safety reporting as it is now will create a lot of unnecessary concern.

Comments from the reviewers:

*** Reviewer #1:

Statistical review

This paper reports a RCT comparing two regimens of chemoprevention vs standard of care for preventing malaria in children with sickle cell anaemia. The trial did not demonstrate differences in the primary endpoint but did find some significant differences in secondary endpoints.

Generally the trial was well-reported and used appropriate statistical methods. I have some minor comments.

1. Abstract: "incidence rates were similar" - I would recommend changing this to "not significantly different" as the confidence intervals allow for fairly large differences in the IRR.

2. Abstract (and throughout results): 'p<0.01' - I would recommend reporting p-values more precisely unless <0.0001.

3. Abstract 'Serious adverse events were common but distributed between groups' - could this be more precisely written that 'distributed between groups', e.g. 'but no arm showed safety concerns'?

4. Abstract: I note the clinicaltrials.gov page has many secondary endpoints that are not mentioned in the abstract. I would recommend (if the abstract reports secondary endpoints) it is clear there were other non-reported secondary endpoints that were not significant.

5. Methods, randomisation: was a fixed block allocation used, or was this stratified or using random block sizes? Providing the block size would be useful.

6. Methods, outcome measures: I would recommend each outcome on the clinicaltrials.gov page is mentioned here and reported in the paper, or a reason for it/them not being reported given here.

7. Methods, statistical analysis: "assuming that each experimental arm would be made" - I would change this to 'each comparison between experimental arm and control would be tested at the 0.0269 level (Dunnett correction)'.

8. Methods, statistical analysis: I did not quite follow how the as-treated analysis worked. Was the indicator for SP-AQ arm a time varying covariate in the model? Later on it is said the AT population was not evaluated - is this just for secondary outcomes?

9. Methods, statistical analysis: I note the protocol said different count outcome analyses would be considered with the one with the lowest residual variance used - this may be worth briefly mentioning in the analysis methods.

10. Results, lines 238-242: similar comment to comment 1.

James Wason

*** Reviewer #2:

This trial compared three drug regimens for malaria chemoprevention (prophylaxis) in patient with sickle cell disease in

western Kenya: daily proguanil (the current recommendation in many countries), monthly SPAQ, and monthly DP.

Unfortunately (from the point of view of the trial aims), there was very little malaria in the trial cohorts,

9 malaria episodes in total. So there was no power to measure a difference in protection against clinical malaria

between the drug regimens. There were more episodes of (asymptomatic) malaria infection, but far fewer than the

sample size calculation envisaged. The main interest will therefore be in the information on tolerability and safety,

but these need to be more clearly presented. The tables need to be revised, they should include the number of each

type of event, and the amount of person time. And there is a problem with the analysis approach in that the conclusions

relate to a comparison of the two monthly regimens, SPAQ and DP, but the statistical analyses are limited to comparison

of each with the proguanil group.

The DSMB stopped the SPAQ arm temporarily, patients in that arm of the trial were switched to proguanil, and then

switched back to SPAQ later, I couldn't see any explanation of the reason for this.

It will be important to explain clearly how much person time was spent on each treatment and the number of doses received of each type of treatment,

I didn't see these details.

I felt that the conclusions were not supported by the data. The fact that the analyses didn't directly compare the SPAQ and DP groups may possibly have led to some confusion about how to interpret the results. For example, referring to SPAQ: "There was no signal of efficacy on parasitologic or hematologic outcomes" and "monthly DP did significantly reduce the incidence of asymptomatic parasitization... and we observed a significant

reduction in dactylitis in the children receiving monthly DP". Such conclusions comparing SPAQ and DP are based on whether or not there was a statistically significant reduction (P<0.05) compared to proguanil, rather than a comparison of the magnitude of reduction between SPAQ and DP.

A closed testing procedure could be have been used for example. (Using the closed testing procedure, first test the global hypothesis with

a 2df level α test, and if it is rejected, then test each of pairwise comparison also at level α.)

It seems unlikely there is any evidence of real differences between SPAQ and DP in this trial. For example, the rate ratio comparing the incidence of dactylis (blocked blood flow in the hands and feet causing painful swelling) in the DP group with the proguanil group was 0.47 (95%CI 0.23 - 0.96), and for the SPAQ group compared to proguanil, 0.78 (95%CI 0.41 - 1.50), it seems unlikely there could be a significant difference between these effects.

And the rate ratios for asymptomatic malaria of 0.46 (0.17 - 1.20) and 0.21 (0.08 - 0.56), also look similar (the fact that the P-value is significant for DP and not for SPAQ does not indicate there is a difference in magnitude of effect).

The conclusion that "Monthly SP-AQ does not appear to have a role as chemoprevention in children with sickle cell anemia" does not seem justified on the basis of these data. There were 10 deaths, 7 of them in the SPAQ arm. The hazard ratio comparing the SPAQ group with the proguanil group had a confidence interval from 0.92 to 32. It seems likely that the cause of death was linked to sickle cell disease rather than drug toxicity. Many millions of children, including many thousands with sickle cell disease, receive SPAQ each year in Seasonal Malaria Chemoprevention programmes. An earlier trial of SPAQ for malaria prevention in sickle cell disease (J Infect Dis. 2015 Aug 15;212(4):617-25) did not find any excess mortality -

that trial, in Nigeria, had 3 arms with a similar number of patients per arm as the current study, there were 7 deaths in total, none of them in the SPAQ group.

A more minor point related to the statistical analysis is the use of Fine & 203 Gray's method to allow death to be a competing risk, this is questionable. It is better to simply treat all competing events as though the individual were right censored at the time the competing event occurs. See https://statisticalhorizons.com/for-causal-analysis-of-competing-risks/

The sample size section mentioned anticipated incidence rate of malaria of 3.7 episodes per person year but then says the study is powered to detect a reduction of 40% with DP and of 40% with proguanil, as if compared to placebo, but there was no placebo group.

The study is potentially interesting but the analyses and interpretation need to be revised.

*** Reviewer #3:

In this manuscript Taylor and colleagues report on the efficacy of daily proguanil versus monthly sulfadoxine-pyrimethamine-amodiaquine (SP-AQ) combination versus monthly dihydroartemisinin-piperaquine (DP) for malaria chemoprevention among Kenyan children with sickle cell disease. This is a very important study as it seeks to evaluate alternative drugs for preventing malaria in one of the groups most vulnerable to malaria and its associated morbidity and mortality. Although the authors found no significant difference in the rate of clinical malaria among children who received monthly SP-AQ or monthly DP compared to those who received daily proguanil possibly due to few clinical malaria cases during follow-up because of indoor residual spraying in the study area, monthly DP was safe and was associated with a lower risk of asymptomatic P. falciparum parasitemia and lower rate of dactylitis. Compared to daily proguanil, monthly SP-AQ was associated with a trend towards an increased risk of mortality. This was a well-conducted study but had two main limitations; the lower than expected clinical malaria episodes which reduced the power of the study and the unanticipated crossover where children randomized to SP-AQ received daily proguanil for about 8 months. It would be useful to the readers if the authors mentioned the cause of the cross-over, which in this manuscript is implied as safety concern, but it is not clear.

Other comments

1. Although there was no significant difference in the risk of clinical malaria comparing AS-AQ and DP to daily proguanil, I would suggest that the authors keep this as the lead finding and conclusion and then highlight a few positive related secondary outcomes e.g risk reduction for asymptomatic parasitemia.

2. In the results tables, it would be useful to clearly indicate numerator and denominator numbers used to estimate the incidence/risk, ie number of episodes of clinical malaria and total number of person years of follow-up for each chemoprevention arm.

Some of the numbers in the tables need to be revised to correct minor errors in rounding off, for example the proportion of households with IRS in table 1 is 26.4%, proportion of children reporting no prior painful crises in the last 12 months is 15.0%. The authors can check all numbers in the tables to ensure accuracy.

3. Line 320, and 359 mention acceptability of DP but the methods of how this was assessed and the data to support this are missing.

*** Reviewer #4:

Monthly sulfadoxine/pyrimethamine-amodiaquine or Dihydroartemisinin-piperaquine as malaria chemoprevention in young children with sickle cell anemia: a randomized controlled trial

Based on the rationale that there is paucity of high-quality comparative effectiveness studies of malaria chemoprevention regimens in African children with sickle cell anaemia (SCA), Taylor and colleagues report results of a trial to evaluate the efficacy of monthly sulfadoxine/pyrimethamine-amodiaquine (SPAQ), or monthly dihydroartemisinin-piperaquine (DP), with daily proguanil as the standard of care, for chemoprevention in children with SCA. The authors report comparable cumulative incidence of clinical malaria and cumulative incidence of painful events as primary and secondary outcome parameters, but slightly lower incidence of dactylitis and P. falciparum parasitaemia in the DP-assigned group.

The authors, in the introduction and rationale, rightly refer to the absence of data on the comparative effect of antimalarial chemoprophylactic regimens as well as lack of consensus on the optimal chemoprophylactic regimen in SCA, which is a population with poor malaria outcomes in endemic areas and in which chemoprevention is important. In this respect, the study by Taylor et. al., is potentially welcome and important contribution to the field.

However, while preventive efficacy of chemoprophylactic agents, the main objective(s) of this study is justified, the safety of repeated antimalarial drug administration specifically in such a population is - I dare say - no less important. From this perspective, several opportunities of this study to demonstrate data that could fill in some of the pressing gaps in knowledge on the (critical) questions surrounding safety of repeated antimalarial drug administration in children in endemic areas in general, and in SCA children in particular, may have been lost.

1. Provide more details on the study site "routine SCA clinic" from where participants were recruited and in addition, include details on the routine SCA care practices (including use of folic acid etc) at the site.

2. Provide a rationale for performing certain investigations (e.g., repeat ECG) in only a selected subset of DP recipients (and why not across the three groups)

3. Per the manuscript (page 9) complete blood count was done every three months. It is surprising that authors present data for only hemoglobin, especially where other parameters (total WBC, platelets) are known to exhibit distinct characteristics in SCA and are linked to the pathophysiology of VOC/painful events as well as other significant SCA complications (e.g., stroke; transfusion requirements etc). Furthermore, the (historical) relationship between amodiaquine and neutropenia/agranulocytosis especially within the context of prophylaxis, is also well known. Given the above, I would strongly encourage authors to show the data for these other hematological parameters as this will strengthen the manuscript along both the safety and pathophysiology domains. In this respect, authors should present not only aggregated summaries but also a) changes from baseline; b) individual outlier data; as well as c) relationships (if any) between the data and the VOC/dactylitis outcome measure(s).

4. Authors should provide more complete information on participants in the "unanticipated crossover" group (number of participants, reasons for the intervention switch etc); suggest to (indeed required, per CONSORT Guidelines) to indicate such a 'flow' in the Trial Profile

5. There is a discrepancy in the number of children in whom followup ECG was conducted (see pages 9 vs 16).

6. The authors report more than 50% incidence of QTc prolongation in (an albeit limited) number of DP recipients. This is an important safety signal that should not, in my opinion, be ignored especially given the known relationship between DP and QTc interval prolongation. Importantly, the statement "given the observed safety of DP….."in the discussion (page 17) is not factually accurate.

7. It is premature to conclude on the safety of DP for prophylaxis in SCA children given the findings and the discussion should be substantially revised to take this into consideration.

8. The summary statement that "our results provide a rationale for the wider use of DP with appropriate monitoring as a routine component of care for children with SCA in malaria endemic settings" is not supported by the data.

9. Other comment: kindly use appropriate classification nomenclature for dactylitis (?hematologic event)

***

Any attachments provided with reviews can be seen via the following link:

[LINK]

PLoS Med. 2022 Oct 10;19(10):e1004104. doi: 10.1371/journal.pmed.1004104.r003

Author response to Decision Letter 1

18 May 2022

Attachment

Submitted filename: EPiTOMISE PlosMed RevResponse final.docx

Click here for additional data file.^{(85.1KB, docx)}

PLoS Med. doi: 10.1371/journal.pmed.1004104.r004

Decision Letter 2

Richard Turner

14 Jun 2022

Dear Dr. Taylor,

Thank you very much for submitting your revised manuscript "Monthly sulfadoxine/pyrimethamine-amodiaquine or dihydroartemisinin-piperaquine as malaria chemoprevention in young Kenyan children with sickle cell anemia: A randomized controlled trial" (PMEDICINE-D-22-00862R2) for consideration at PLOS Medicine.

Your paper was discussed with our academic editor and re-seen by three independent reviewers, including our statistical reviewer. The reviews are appended at the bottom of this email and any accompanying reviewer attachments can be seen via the link below:

[LINK]

In the light of these reviews, we will again be unable to accept the manuscript for publication in the journal in its current form, but we would like to invite you to submit a further revised version that addresses the reviewers' and editors' comments fully. You will recognize that we cannot make a decision on possible publication until we have seen the revised manuscript and your response, and we may seek re-review by one or more of the reviewers.

We hope to receive your revised manuscript by Jul 08 2022 11:59PM. Please email us (plosmedicine@plos.org) if you have any questions or concerns.

Please use the following link to submit the revised manuscript:

https://www.editorialmanager.com/pmedicine/

Your article can be found in the "Submissions Needing Revision" folder.

Please let me know if you have any questions, and we look forward to receiving your revised manuscript.

Sincerely,

Richard Turner, PhD

Senior editor, PLOS Medicine

rturner@plos.org

-----------------------------------------------------------

Requests from the editors:

As mentioned previously, please adapt the data statement (submission form) so as to comply with PLOS' data policy, https://journals.plos.org/plosmedicine/s/data-availability.

Noting the comments from one referee, please report additional measures as appropriate to their inclusion in the study protocol/SAP (i.e., identifying comparisons as post-hoc if needed).

Please add a few words to the abstract to identify the trial site.

Please correct the referencing at lines 383-4.

Throughout the text, please move reference call-outs before punctuation (e.g., "... occlusive complications [6,7]."; noting the absence of spaces within the square brackets).

Noting reference 29 and others, please use the journal name abbreviation "PLoS ONE".

Please correct the spelling of the first author's name for reference 36.

We did not find a completed CONSORT checklist with your revision and ask that you include this with your next revision.

In the checklist, please refer to individual items by section (e.g., 'Methods') and paragraph number, not by line or page numbers as these generally change in the event of publication.

Comments from academic editor:

It may be most efficient to ask the authors to provide an additional direct comparison of

a) Text: Incidence of Pf infections in the DP vs SP-AQ arm

b) Text: Incidence of dactylitis in the DP vs SP-AQ arm

c) Table 1 and 2: add direct comparisons of DP vs SP-AQ arm

That should address “comparisons throughout the paper”.

The authors should review their conclusions (abstract, summary, and discussion) if warranted by the findings.

Comments from the reviewers:

*** Reviewer #1:

Thank you to the authors for addressing my previous comments well. I would still recommend briefly noting the deviation from which endpoints were specified in the SAP compared to ct.gov (perhaps using more diplomatic language than in the response to comment 28!) but will leave that as a suggestion to the editor. Other that that I have no further issues to raise.

*** Reviewer #2:

I think the presentation of results is misleading in the sense that the two interventions are being compared based on

comparing the P-values for the difference of each with control, for example: "The incidence of P. falciparum infection relative to daily

Proguanil was similar in the monthly SP-AQ group (IRR 0.46; 95% CI: 0.17—1.20; p=0.13) but reduced with monthly DP (IRR 0.21; 9 5% CI: 0.08—0.56; p=0.002)." But it seems unlikely that the efficacy differs between the two interventions. Similarly, the rate ratio comparing the incidence of dactylis (blocked blood flow in the hands and feet causing painful swelling) in the DP group with the proguanil group was 0.47 (95%CI 0.23 - 0.96), and for the SPAQ group compared to proguanil, 0.78 (95%CI 0.41 - 1.50), it seems unlikely there could be a statistically significant difference between these effects. The difference in mortality is also likely to be a chance finding. There needs to be a head-on comparison of the two groups. This applies to comparisons throughout the paper.

*** Reviewer #3:

Two minor comments:

1. The primary outcome could also lead the concluding parts of the abstract and the discussion. The authors can first highlight lack of difference in the primary outcome, probably mention the reason why, and then go on to mention important significant secondary outcomes like parasite prevalence, like in the statement below.

"Although there was no significant difference in the incidence of clinical malaria between the three treatment arms, possibly due to reduced malaria transmission intensity, DP was associated with……….then go on to mention important secondary outcomes that were significant."

2. Line 371, I suggest that "acceptability" be removed from the statement. To me, data presented in supplemental table 1 supports adherence not acceptability.

***

Any attachments provided with reviews can be seen via the following link:

[LINK]

PLoS Med. 2022 Oct 10;19(10):e1004104. doi: 10.1371/journal.pmed.1004104.r005

Author response to Decision Letter 2

20 Jun 2022

Attachment

Submitted filename: EPiTOMISE PlosMed RevReResponse.docx

Click here for additional data file.^{(70.1KB, docx)}

PLoS Med. doi: 10.1371/journal.pmed.1004104.r006

Decision Letter 3

Caitlin Moyer

10 Aug 2022

Dear Dr. Taylor,

Thank you very much for re-submitting your manuscript "Monthly sulfadoxine/pyrimethamine-amodiaquine or dihydroartemisinin-piperaquine as malaria chemoprevention in young Kenyan children with sickle cell anemia: A randomized controlled trial" (PMEDICINE-D-22-00862R3) for review by PLOS Medicine.

I have discussed the paper with my colleagues and the academic editor and it was also seen again by one reviewer. I am pleased to say that provided the remaining editorial and production issues are dealt with we are planning to accept the paper for publication in the journal.

The remaining issues that need to be addressed are listed at the end of this email. Any accompanying reviewer attachments can be seen via the link below. Please take these into account before resubmitting your manuscript:

[LINK]

In revising the manuscript for further consideration here, please ensure you address the specific points made by each reviewer and the editors. In your rebuttal letter you should indicate your response to the reviewers' and editors' comments and the changes you have made in the manuscript. Please submit a clean version of the paper as the main article file. A version with changes marked must also be uploaded as a marked up manuscript file.

Please also check the guidelines for revised papers at http://journals.plos.org/plosmedicine/s/revising-your-manuscript for any that apply to your paper. If you haven't already, we ask that you provide a short, non-technical Author Summary of your research to make findings accessible to a wide audience that includes both scientists and non-scientists. The Author Summary should immediately follow the Abstract in your revised manuscript. This text is subject to editorial change and should be distinct from the scientific abstract.

We expect to receive your revised manuscript within 1 week. Please email us (plosmedicine@plos.org) if you have any questions or concerns.

We ask every co-author listed on the manuscript to fill in a contributing author statement. If any of the co-authors have not filled in the statement, we will remind them to do so when the paper is revised. If all statements are not completed in a timely fashion this could hold up the re-review process. Should there be a problem getting one of your co-authors to fill in a statement we will be in contact. YOU MUST NOT ADD OR REMOVE AUTHORS UNLESS YOU HAVE ALERTED THE EDITOR HANDLING THE MANUSCRIPT TO THE CHANGE AND THEY SPECIFICALLY HAVE AGREED TO IT.

Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript.

Please note, when your manuscript is accepted, an uncorrected proof of your manuscript will be published online ahead of the final version, unless you've already opted out via the online submission form. If, for any reason, you do not want an earlier version of your manuscript published online or are unsure if you have already indicated as such, please let the journal staff know immediately at plosmedicine@plos.org.

If you have any questions in the meantime, please contact me or the journal staff on plosmedicine@plos.org.

We look forward to receiving the revised manuscript by Aug 17 2022 11:59PM.

Sincerely,

Caitlin Moyer, PhD

Associate Editor

PLOS Medicine

plosmedicine.org

------------------------------------------------------------

Requests from Editors:

1. Data availability statement: The Data Availability Statement (DAS) requires revision. Please spell out “ICF” in the statement rather than using an acronym. It would be helpful to include a general web address where interested researchers can learn more about the dataset/ accessing the data. Please note that a study author cannot be the contact person for the data.

2. Funding statement: Please add the following information to the funding statement: “SP-AQ was supplied free of charge to the trial by Guilin Pharmaceuticals, which had no role in the design, conduct, analysis, reporting, or decision to report the results.”

3. Throughout: Please be sure that all abbreviations and acronyms are defined in the text at their first point of use (both for the Abstract and the main text).

4. Abstract: Background: We suggest an additional sentence providing context on the current chemoprevention practices or similar.

5. Abstract: Line 40: Please define HbSS at first use in the text. Please describe some brief information on the inclusion criteria/recruitment of the children with HbSS. Please note in the abstract that children were followed up monthly for 12 months.

6. Abstract: Line 45: Please provide a brief description of painful events and how this outcome was determined. Please list other secondary outcomes, including: parasitologic outcomes and hematologic outcomes.

7. Abstract: Line 58-59: Please point out that the differences in numbers of deaths between groups did not reach statistical significance.

8. Abstract: Conclusions: We suggest revising the first sentence to address the study implications without overreaching what can be concluded from the data, commenting on the primary outcome first and then key secondary findings; the phrase "In this study, we observed ..." may be useful. We suggest: In this study, we observed that monthly malaria chemoprevention with monthly SP-AQ or monthly DP did not significantly reduce incidence of clinical malaria compared with daily Proguanil. Monthly DP was associated with a lower rate of dactylitis and infection with P. falciparum relative to the Proguanil group, despite limited malaria transmission during the study period.” Please note that there is usually a distinction in the language in terms of causal vs. associational for primary and secondary trial outcomes. It would be beneficial to use associational language throughout when describing secondary outcomes.

9. Author summary: Line 93-94: Please rephrase slightly to: “...a higher rate of deaths occured in the SP-AQ group, but this did not reach statistical significance.” or similar.

10. Author summary: Line 98-99: Please revise to: “The observed greater number of deaths among children with SCA in the SP-AQ treated group was unexpected, and while this association did not reach statistical significance, further evaluation is warranted.” or similar.

11. Methods: Line 145-146: Please revise to: “"This study is reported as per the Consolidated Standards of Reporting Trials (CONSORT) guideline (S1 Checklist)."

12. Methods: Please provide additional details on eligibility in the main text of the Methods.

13. Methods: Lines 196 and 209: Please make sure that a complete listing of all primary and secondary outcomes, and safety outcomes, is included here.

14. Methods: Line 223-229: Please provide an explanation for the cross-over from SP-AQ to Proguanil.

15. Methods: Line 242-244: Please describe changes to the protocol, e.g. those prompted by the pandemic, in the text. Please mention and explain the reasons for the modifications including: exclusion of children with low hemoglobin, the collection of baseline laboratory testing at screening rather than enrollment, and the enhanced monthly hemoglobin monitoring, SAE review, and patient referral. Please complete and include the CONSERVE checklist, if relevant (https://www.equator-network.org/reporting-guidelines/guidelines-for-reporting-trial-protocols-and-completed-trials-modified-due-to-the-covid-19-pandemic-and-other-extenuating-circumstances-the-conserve-2021-statement/)

16. Results: Line 302: Please revise to: “...DP was associated with reduced rates of asymptomatic parasitization…”

17. Results: Line 315: Please revise to: “...did not modify the association between DP and dactylitis…”

18. Results: Please report p values in the text throughout, where appropriate, for example at lines 318-333.

19. Results: Lines 329-333: Please revise to indicate that the observed increase in risk of death among SP-AQ recipients (compared with the Proguanil or with the DP treated groups) was not statistically significant.

20. Discussion: Line 357-358: Please revise to: “...monthly DP was associated with significantly lower incidence of asymptomatic parasitization.”

21. Discussion: Line 365: Please revise to: “Monthly DP was associated with a reduced rate of episodes of dactylitis compared to daily Proguanil…”

22. Discussion: Line 398-399: It is not clear that the data support this statement, and we suggest removing it.

23. Discussion: line 426-427: Please mention the primary study outcome up front in the Conclusion paragraph. Please revise the sentence to: “...monthly DP was observed to be acceptable and was associated with reduced incidence of dactylitis and asymptomatic P. falciparum infection…”

24. Discussion: Line 429: We suggest removing: “and was associated with unacceptable morbidity.”

25. Figure 1: Please define all abbreviations used in the legend.

26. Figure 2: Please define all abbreviations used in the legend. On the axis, please make it clear that the point estimates are relative for each SP-AQ or DP relative to Proguanil, not relative to each other.

27. Table 1: It seems as though p values should not be needed as participants were randomized.

28. Table 3: Where reporting the p value for asymptomatic P. falciparum infection, please do not report as p<0.01. Please report the exact p value, unless p<0.001.

29. Reference 29, 34, and 36: Please change the journal to PLoS One.

30. CONSORT checklist: Thank you for including the CONSORT checklist. On the checklist, please make it clear you are referring to paragraph numbers within sections. For “Protocol” please report the location as Supporting Information files, or similar. For “Funding” please report the location as Funding statement, or similar.

31. Study Protocol: Thank you for including a copy of the study protocol. Please confirm that the image used in Appendix A-E is not reproduced from other sources that are not CC-BY. Please see https://journals.plos.org/plosmedicine/s/figures#loc-licenses-and-copyright for more information.

32. Supplemental Methods: Please move the description of inclusion/exclusion criteria and sample size considerations to the main text.

33. Supplemental Figure: Please define all abbreviations used in a descriptive legend.

34. Supporting information Table S6 and S7: Please define “-” and “(-,-)” in the legend.

35. Supporting information: Please check reference list formatting. Please use the "Vancouver" style for reference formatting, and see our website for other reference guidelines https://journals.plos.org/plosmedicine/s/submission-guidelines#loc-references

Comments from Reviewers:

Reviewer #5: This is a well written paper.

The authors have adequately addressed the comments made by the reviewers.

I would suggest adding a column for p-values in Table 1.

Any attachments provided with reviews can be seen via the following link:

[LINK]

PLoS Med. 2022 Oct 10;19(10):e1004104. doi: 10.1371/journal.pmed.1004104.r007

Author response to Decision Letter 3

24 Aug 2022

Attachment

Submitted filename: EPiTOMISE PlosMed 3rdResubResponse.docx

Click here for additional data file.^{(73.5KB, docx)}

PLoS Med. doi: 10.1371/journal.pmed.1004104.r008

Decision Letter 4

Caitlin Moyer

26 Aug 2022

Dear Dr Taylor,

On behalf of my colleagues and the Academic Editor, Lorenz von Seidlein, I am pleased to inform you that we have agreed to publish your manuscript "Monthly sulfadoxine/pyrimethamine-amodiaquine or dihydroartemisinin-piperaquine as malaria chemoprevention in young Kenyan children with sickle cell anemia: A randomized controlled trial" (PMEDICINE-D-22-00862R4) in PLOS Medicine.

Before your manuscript can be formally accepted you will need to complete some formatting changes, which you will receive in a follow up email. Please be aware that it may take several days for you to receive this email; during this time no action is required by you. Once you have received these formatting requests, please note that your manuscript will not be scheduled for publication until you have made the required changes.

In the meantime, please log into Editorial Manager at http://www.editorialmanager.com/pmedicine/, click the "Update My Information" link at the top of the page, and update your user information to ensure an efficient production process.

Please also address the following editorial requests:

-Abstract: Lines 60-63: Please revise to: “Serious adverse events were common and distributed between groups, though compared to daily Proguanil (n=2), more children died receiving monthly SP-AQ (n=7; Hazard Ratio [HR] 5.44; 95% CI: 0.92—32.11) but not DP (n=1; HR 0.61; 95% CI 0.04—9.22), though differences did not reach statistical significance for either SP-AQ or DP.” Please also add p values for these results.

-Results: Line 356-360: Please revise to: “Ten children died: 2 receiving daily Proguanil, 7 receiving SP-AQ, and 1 receiving DP (Table 4); relative to Proguanil (cumulative IR [CIR] 2.2%), the risk of death was elevated in SP-AQ recipients, though this did not reach statistical significance (CIR 11.3%; Hazard Ratio [HR] 5.44; 95% CI 0.92 – 32.11; p=0.064), and was not elevated in DP recipients (CIR 1.3%; HR 0.61; 95% CI 0.04 – 9.22; p=0.89).

Discussion: Lines 385-387: Please revise to: “Compared to Proguanil, neither monthly SP-AQ nor DP reduced the incidence of the primary outcome clinical malaria, though monthly DP did significantly reduce the incidence of asymptomatic parasitization.”

-Discussion: Lines 458-460: Please revise to: “Monthly SP-AQ had no measurable benefit compared to daily Proguanil, and we observed a greater number of deaths in this group, though mortality rate differences were not statistically significant.”

-Figure 1: In the legend, please define QTcF, Hb, ALT, LAR.

-Study Protocol: Please remove or replace any image (e.g. in the Study Protocol) for which the source and license cannot be determined. Please see https://journals.plos.org/plosmedicine/s/licenses-and-copyright for further information.

-References:

--Reference 2: Please abbreviate as Lancet Glob Health

--Reference 19: Please abbreviate as Lancet Haematol

--Reference 6 and 25: Please abbreviate as Br Med J

PRESS

We frequently collaborate with press offices. If your institution or institutions have a press office, please notify them about your upcoming paper at this point, to enable them to help maximise its impact. If the press office is planning to promote your findings, we would be grateful if they could coordinate with medicinepress@plos.org. If you have not yet opted out of the early version process, we ask that you notify us immediately of any press plans so that we may do so on your behalf.

We also ask that you take this opportunity to read our Embargo Policy regarding the discussion, promotion and media coverage of work that is yet to be published by PLOS. As your manuscript is not yet published, it is bound by the conditions of our Embargo Policy. Please be aware that this policy is in place both to ensure that any press coverage of your article is fully substantiated and to provide a direct link between such coverage and the published work. For full details of our Embargo Policy, please visit http://www.plos.org/about/media-inquiries/embargo-policy/.

Thank you again for submitting to PLOS Medicine. We look forward to publishing your paper.

Sincerely,

Caitlin Moyer, Ph.D.

Associate Editor

PLOS Medicine

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

S1 CONSORT Checklist. CONSORT checklist.

(DOC)

Click here for additional data file.^{(218KB, doc)}

S1 Protocol. Study protocol.

(DOCX)

Click here for additional data file.^{(711.2KB, docx)}

S1 Analysis. Statistical analysis plan.

(DOCX)

Click here for additional data file.^{(54.6KB, docx)}

S1 Text. Supporting information.

Supplemental methods, figure, and tables.

(DOCX)

Click here for additional data file.^{(98.4KB, docx)}

Attachment

Submitted filename: EPiTOMISE PlosMed RevResponse final.docx

Click here for additional data file.^{(85.1KB, docx)}

Attachment

Submitted filename: EPiTOMISE PlosMed RevReResponse.docx

Click here for additional data file.^{(70.1KB, docx)}

Attachment

Submitted filename: EPiTOMISE PlosMed 3rdResubResponse.docx

Click here for additional data file.^{(73.5KB, docx)}

Data Availability Statement

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PERMALINK

Monthly sulfadoxine/pyrimethamine-amodiaquine or dihydroartemisinin-piperaquine as malaria chemoprevention in young Kenyan children with sickle cell anemia: A randomized controlled trial

Steve M Taylor

Sarah Korwa

Angie Wu

Cynthia L Green

Betsy Freedman

Sheila Clapp

Joseph Kipkoech Kirui

Wendy P O’Meara

Festus M Njuguna

Roles

Abstract

Background

Methods and findings

Conclusions

Trial registration

Author summary

Why was this study done?

What did the researchers do and find?

What do these findings mean?

Introduction

Methods

Trial design and oversight

Trial setting and population

Trial interventions

Trial assessments

Outcome measures

Statistical analysis

Results

Participants and follow-up

Fig 1. Screening, randomization, enrollment, and follow-up.

Table 1. Characteristics of the patients at randomization.

Efficacy

Table 2. Main parasitologic and hematologic outcomes.

Table 3. Secondary outcomes in the AT population.

Fig 2. Main hematologic outcomes in the AT population according to subgroup.

Safety

Table 4. Safety outcomes and SAEs in ATP population.

Discussion

Supporting information

Acknowledgments

Abbreviations

Data Availability

Funding Statement

References

Decision Letter 0

Richard Turner

Roles

Decision Letter 1

Richard Turner

Roles

Author response to Decision Letter 1

Decision Letter 2

Richard Turner

Roles

Author response to Decision Letter 2

Decision Letter 3

Caitlin Moyer

Roles

Author response to Decision Letter 3

Decision Letter 4

Caitlin Moyer

Roles

Associated Data

Supplementary Materials

Data Availability Statement

ACTIONS

PERMALINK

RESOURCES

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